Method for retrovirus removal

Abstract

A method for removing retroviruses from liquid samples and a nanofiber containing liquid filtration medium that simultaneously exhibits high liquid permeability and high microorganism retention is disclosed. Retroviruses are removed from a liquid by passing the liquid through a porous nanofiber containing filtration medium having a retrovirus LRV greater than about 6, and the nanofiber(s) has a diameter from about 10 nm to about 100 nm. The filtration medium can be in the form of a fibrous electrospun polymeric nanofiber liquid filtration medium mat.

Claims

1. A porous nanofiber containing filtration medium comprising a porous nanofiber mat made by electrospinning a polymer, wherein the porous nanofiber mat has a retrovirus Log Reduction Value (LRV) greater than about 6, a porosity from about 80% to about 95%, and a liquid permeability greater than about 100 LMH/psi, and wherein the nanofiber mat has a fiber diameter effective for retention of retroviruses by size-based exclusion.

2. The porous nanofiber containing filtration medium of claim 1, wherein the porous nanofiber mat exhibits a mean flow bubble point, as tested with isopropanol, from 100 to 150 psi.

3. The porous nanofiber containing filtration medium of claim 2, wherein the porous nanofiber mat has a fiber diameter from about 25-30 nm.

4. The porous nanofiber containing filtration medium of claim 2, wherein the porous nanofiber mat has a fiber diameter from about 10-100 nm.

5. The porous nanofiber containing filtration medium of claim 2, wherein the porous nanofiber mat has a liquid permeability of at least about 300 LMH/psi.

6. The porous nanofiber containing filtration medium of claim 1, wherein the filtration medium has a thickness from about 1 ?m to about 500 ?m.

7. The porous nanofiber containing filtration medium of claim 1, wherein the porous nanofiber mat has a thickness from 1 ?m to 50 ?m.

8. The porous nanofiber containing filtration medium of claim 7, wherein the porous nanofiber mat has a fiber diameter from about 25-30 nm.

9. The porous nanofiber containing filtration medium of claim 7, wherein the porous nanofiber mat has a fiber diameter from about 10-100 nm.

10. The porous nanofiber containing filtration medium of claim 7, wherein the porous nanofiber mat has a liquid permeability of at least about 300 LMH/psi.

11. The porous nanofiber containing filtration medium of claim 1, wherein the retrovirus is Human Immunodeficiency Virus (HIV).

12. The porous nanofiber containing filtration medium of claim 1, wherein the retrovirus is Human T-cell leukemia virus (HTLV).

13. The porous nanofiber containing filtration medium of claim 1, wherein the polymer is selected from the group consisting of polyimide, aliphatic polyamide, aromatic polyamide, polysulfone, cellulose acetate, polyether sulfone, polyurethane, poly(urea urethane), polybenzimidazole, polyetherimide, polyacrylonitrile, poly(ethylene terephthalate), polypropylene, poly aniline, poly(ethylene oxide), poly(ethylene naphthalate), poly(butylene terephthalate), styrene butadiene rubber, polystyrene, poly(vinyl chloride), poly(vinyl alcohol), poly(vinylidene fluoride), poly(vinyl butylene) and copolymers, derivative compounds, or blends thereof.

14. The porous nanofiber containing filtration medium of claim 1, wherein the polymer comprises an aliphatic polyamide.

15. The porous nanofiber containing filtration medium of claim 1, wherein the polymer comprises a blend of polymers or copolymers.

16. The porous nanofiber containing filtration medium of claim 1, wherein the porous nanofiber mat is disposed on a porous support.

17. The porous nanofiber containing filtration medium of claim 16, wherein the porous support comprises one or more layers selected from the group consisting of spunbonded nonwovens, meltblown nonwovens, needle punched nonwovens, spunlaced nonwovens, wet laid nonwovens, resin-bonded nonwovens, woven fabrics, knit fabrics, paper, and combinations thereof.

18. The porous nanofiber containing filtration medium of claim 1, wherein the porous nanofiber mat has a fiber diameter from about 25-30 nm.

19. The porous nanofiber containing filtration medium of claim 1, wherein the porous nanofiber mat has a fiber diameter from about 10-100 nm.

20. The porous nanofiber containing filtration medium of claim 1, wherein the porous nanofiber mat has a liquid permeability of at least about 300 LMH/psi.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate the presently contemplated embodiments of the invention and, together with the description, serve to explain the principles of the invention.

(2) FIG. 1 is a schematic of the process of electrospinning a nanofiber according to one embodiment of the invention.

(3) FIG. 2 is a frontal scanning electron micrograph of nylon fibers from an embodiment of the invention exemplified in Example 1.

(4) FIG. 3 is a frontal scanning electron micrograph of nylon fibers from an embodiment of the invention exemplified in Example 3.

(5) FIG. 4 is a graph of bubble point distribution for a nylon fiber mat described in Example 1 and for a commercial retrovirus retentive membrane.

(6) FIG. 5 is a graph of Log Reduction Value (LRV) of bacteriophage PR772 as a function of spin time of a nanofiber mat.

DESCRIPTION OF THE EMBODIMENTS

(7) All publications, patents and patent applications cited herein, whether supra or infra, are hereby incorporated by reference in their entirety to the same extent as if each individual publication, patent or patent application was specifically and individually indicated to be incorporated by reference.

(8) For the purposes of this specification and appended claims, unless otherwise indicated, all numbers expressing quantities of ingredients, percentages or proportions of materials, reaction conditions, and other numerical values used in the specification and claims, are to be understood as being modified in all instances by the term about.

(9) Accordingly, unless indicated to the contrary, the numerical parameters set forth in the following specification and attached claims are approximations that may vary depending upon the desired properties sought to be obtained by the present invention. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be construed in light of the number of reported significant digits and by applying ordinary rounding techniques.

(10) Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the invention are approximations, the numerical values set forth in the specific examples are reported as precisely as possible. Any numerical value, however, inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements. Moreover, all ranges disclosed herein are to be understood to encompass all subranges subsumed therein. For example, a range of 1 to 10 includes any and all subranges between (and including) the minimum value of 1 and the maximum value of 10, that is, any and all subranges having a minimum value of equal to or greater than 1 and a maximum value of equal to or less than 10, e.g., 5.5 to 10.

(11) Before describing the present invention in further detail, a number of terms will be defined. Use of these terms does not limit the scope of the invention but only serve to facilitate the description of the invention.

(12) As used herein, the singular forms a, an, and the include plural referents unless the context clearly dictates otherwise.

(13) The term nanofibers refers to fibers having diameters varying from a few tens of nanometers up to several hundred nanometers, but generally less than one micrometer.

(14) The terms filter medium or filter media refer to a material, or collection of material, through which a fluid carrying a microorganism contaminant passes, wherein microorganism is deposited in or on the material or collection of material.

(15) The terms flux and flow rate are used interchangeably to refer to the rate at which a volume of fluid passes through a filtration medium of a given area.

(16) The filtration medium of the present invention includes a porous electrospun nanofiber liquid filtration mat. The nanofibers have an average fiber diameter of about 10 nm to about 100 nm. The filtration medium has a mean pore size ranging from about 0.05 um to about 1 um. The filtration medium has a porosity ranging from about 80% to about 95%. The filtration medium has a thickness ranging from about 1 ?m to about 500 ?m, preferably from about 10 ?m and about 100 ?m. The filtration medium has liquid permeability greater than about 100 LMH/psi.

(17) Polymers suitable for use in the nanofibers of the invention include thermoplastic and thermosetting polymers. Suitable polymers include, but are not limited to, nylon, polyimide, aliphatic polyamide, aromatic polyamide, polysulfone, cellulose, cellulose acetate, polyether sulfone, polyurethane, poly(urea urethane), polybenzimidazole, polyetherimide, polyacrylonitrile, poly(ethylene terephthalate), polypropylene, polyaniline, poly(ethylene oxide), poly(ethylene naphthalate), poly(butylene terephthalate), styrene butadiene rubber, polystyrene, poly(vinyl chloride), poly(vinyl alcohol), poly(vinylidene fluoride), poly(vinyl butylene), copolymers, derivative compounds and blends thereof, and combinations thereof.

(18) The process for making the electrospun nanofiber mat of the filtration medium is disclosed in WO 2005/024101, WO 2006/131081, and WO 2008/106903, each fully incorporated herein by reference, and each assigned to Elmarco S. R. O., of Liberec, Czech Republic.

(19) In one embodiment of the present invention as depicted in FIG. 1, the filtration medium comprises a porous mat 60 made from a single nanofiber, wherein the single nanofiber is made by a single pass of a moving collection apparatus 30 positioned between a rotating drum 20 and a collector 35 through the electrospinning process. It will be appreciated that the fibrous web can be formed by one or more spinning drums 20 running simultaneously above the same moving collection apparatus 30.

(20) In FIG. 1, the moving collection apparatus 30 is preferably a moving collection belt positioned within an electrostatic field 50 between the spinning drum 20 and the collector 35, wherein a polymer solution 10 can be subjected to an electrical potential from a high voltage source 40, such that an electrospun polymer fiber is produced in the electric field 50.

(21) In one embodiment of the invention, a fibrous mat is made by depositing nanofiber (s) from a nylon solution. The nanofiber mat has a basis weight of between about 1 g/m.sup.2 and about 10 g/m.sup.2, as measured on a dry basis, i.e., after the residual solvent has evaporated or been removed.

(22) In one embodiment of the invention, any of a variety of porous single or multilayered substrates or supports can be arranged on a moving collection belt to collect and combine with the electrospun nanofiber mat medium, forming a composite filtration device.

(23) Examples of single or multilayered porous substrates or supports include, but are not limited to, spunbonded nonwovens, meltblown nonwovens, needle punched nonwovens, spunlaced nonwovens, wet laid nonwovens, resin-bonded nonwovens, woven fabrics, knit fabrics, paper, and combinations thereof.

(24) In another embodiment of the invention the electrospun nanofiber mat medium taught herein may be bonded to a porous substrate or support. Bonding may be accomplished by known methods in the art, including but not limited to thermal calendaring between heated smooth nip rolls, ultrasonic bonding, and through gas bonding. Bonding the electrospun nanofiber medium to a support increases the strength and the compression resistance of the medium, such that the medium may withstand the forces associated with being handled such as when the porous medium is formed into a useful filter and/or installed into a filtration device. In addition, the physical properties of the porous electrospun nanofiber medium such as thickness, density, and the size and shape of the pores may be affected depending on the bonding methods used.

(25) For instance, thermal calendering can be used to reduce the thickness and increase the density and reduce the porosity of the electrospun nanofiber mat medium, and reduce the size of the pores. This in turn decreases the flow rate through the medium at a given applied differential pressure.

(26) In general, ultrasonic bonding will bond to a smaller area of the porous electrospun nanofiber medium than thermal calendering, and therefore has a lesser effect on thickness, density and pore size.

(27) Gas bonding generally has minimal effect on the thickness, density and pore size of the porous electrospun nanofiber medium, therefore this bonding method may be preferable in applications in which maintaining higher fluid flow rate is desired.

(28) When thermal calendering is used, care must be taken not to over-bond the porous electrospun nanofiber material, such that the nanofibers melt and no longer retain their structure as individual fibers. In the extreme, over-bonding would result in the nanofibers melting completely such that a film would be formed. One or both of the nip rolls used is heated to a temperature of between about ambient temperature, e.g., about 25? C. and about 300? C. The porous nanofiber medium and/or porous support or substrate, can be compressed between the nip rolls at a pressure ranging from about 0 lb/in to about 1000 lb/in (178 kg/cm). The porous nanofiber medium can be compressed at a line speed of at least about 10 ft/min (3 m/min).

(29) Calendering conditions, e.g., roll temperature, nip pressure and line speed, can be adjusted to achieve the desired solidity. In general, application of higher temperature, pressure, and/or residence time under elevated temperature and/or pressure results in increased solidity.

(30) Other mechanical steps, such as stretching, cooling, heating, sintering, annealing, reeling, unreeling, and the like, may optionally be included in the overall process of forming, shaping and making the electrospun nanofiber mat medium as desired.

(31) For example, the electrospun nanofiber mat medium taught herein may be stretched in a single step or a plurality of steps as desired. Depending on the stretching method used to stretch the electrospun nanofiber mat medium, stretching can adjust the physical properties of the mat including thickness, density, and the size and shape of the pores formed in the mat. For example, if the electrospun nanofiber mat is stretched in a single direction (uniaxial stretching), the stretching may be accomplished by a single stretching step or a sequence of stretching steps until the desired final stretch ratio is attained.

(32) Similarly, if the electrospun nanofiber mat medium is stretched in two directions (biaxial stretching), the stretching can be conducted by a single biaxial stretching step or a sequence of biaxial stretching steps until the desired final stretch ratios are attained. Biaxial stretching may also be accomplished by a sequence of one or more uniaxial stretching steps in one direction and one or more uniaxial stretching steps in another direction. Biaxial stretching steps where the electrospun nanofiber mat is stretched simultaneously in two directions and uniaxial stretching steps may be conducted in sequence in any order.

(33) Methods for stretching the mat are not particularly limited, and use may be made of ordinary tentering, rolling, or inflation or a combination of two or more of these. The stretching may be conducted uniaxially, biaxially, etc. In the case of biaxial stretching, machine-direction stretching and transverse-direction stretching may be conducted either simultaneously or successively.

(34) Various types of stretching apparatus are well known in art and may be used to accomplish stretching of the electrospun mat according to the present invention. Uniaxial stretching is usually accomplished by stretching between two rollers wherein the second or downstream roller rotates at a greater peripheral speed than the first or upstream roller. Uniaxial stretching can also be accomplished on a standard tentering machine.

(35) Biaxial stretching may be accomplished by simultaneously stretching in two different directions on a tentering machine. More commonly, however, biaxial stretching is accomplished by first uniaxially stretching between two differentially rotating rollers as described above, followed by either uniaxially stretching in a different direction using a tenter machine or by biaxially stretching using a tenter machine. The most common type of biaxial stretching is where the two stretching directions are approximately at right angles to each other. In most cases where a continuous sheet is being stretched, one stretching direction is at least approximately parallel to the long axis of the sheet (machine direction) and the other stretching direction is at least approximately perpendicular to the machine direction and is in the plane of the sheet (transverse direction).

(36) After the electrospun nanofiber mat has been stretched either uniaxially or biaxially, the stretched porous electrospun nanofiber mat can again be calendared. The stretched electrospun nanofiber mat can be forwarded to a pair of heated calendar rolls acting cooperatively so as to form a mat of reduced thickness compared to the mat exiting from the stretching apparatus. By regulating the pressure exerted by these calendar rolls along with the temperature, the pore size of the final electrospun nanofiber mat can be controlled as desired, thereby allowing for the adjustment of the average pore size.

(37) The electrospun nanofiber mat may be heated by any of a wide variety of techniques prior to, during, and/or after stretching. Examples of these techniques include radiative heating such as that provided by electrically heated or gas fired infrared heaters, convective heating such as that provided by recirculating hot air, and conductive heating such as that provided by contact with heated rolls. The temperatures which are measured for temperature control purposes may vary according to the apparatus used and personal preference.

(38) In general, the temperature or temperatures can be controlled such that the electrospun nanofiber mat is stretched about evenly so that the variations, if any, in thickness of the stretched mat are within acceptable limits and so that the amount of stretched microporous electrospun nanofiber mat outside of those limits is acceptably low. It will be apparent that the temperatures used for control purposes may or may not be close to those of the electrospun nanofiber mat itself since they depend upon the nature of the apparatus used, the locations of the temperature-measuring devices, and the identities of the substances or objects whose temperatures are being measured.

(39) The porosity can be modified as a result of calendering. The range of porosity from about 5% to about 90% can be obtained.

(40) While filtration medium is often used in single-layer configuration, it is sometimes advantageous to provide more than one layer of filtration medium adjacent to each other. Layering membrane filters to improve particle retention is commonly used in virus filtration and is practiced commercially in Millipore's product lines of Viresolve? NFP and Viresolve Pro?. Layering filtration media of the same or different composition is also used to improve filter throughput. Examples of such layered filters are Millipore's Express? SHC and SHRP product lines.

(41) Other considerations for choosing a multi-layered filtration product include economics and convenience of media and device manufacturing, ease of sterilization and validation. The fibrous filtration media of the present invention can be used in single-layer or in a multi-layer configuration.

(42) The preferred layer configuration is often selected based on practical considerations. These considerations take into account the known relationship between LRV and thickness, whereby LRV typically increases with thickness. A practitioner can select multiple ways of achieving desired level of LRV, e.g. by using fewer layers of larger thickness or larger number of thinner layers.

(43) Test Methods

(44) Basis Weight was determined by ASTM D-3776, which is incorporated herein by reference and reported in g/m.sup.2.

(45) Porosity was calculated by dividing the basis weight of the sample in g/m.sup.2 by the polymer density in g/cm.sup.3, by the sample thickness in micrometers, multiplying by 100, and subtracting the resulting number from 100, i.e., porosity=100?[basis weight/(density.times.thickness).times.100].

(46) Fiber Diameter was determined as follows. A scanning electron microscope (SEM) image was taken at 60,000 times magnification of each side of nanofiber mat sample. The diameter of ten (10) clearly distinguishable nanofibers were measured from each SEM image and recorded. Defects were not included (i.e., lumps of nanofibers, polymer drops, intersections of nanofibers). The average fiber diameter of both side for each sample was calculated.

(47) Thickness was determined by ASTM D1777-64, which is incorporated herein by reference, and is reported in micrometers.

(48) Mean flow bubble point was measured according to ASTM Designation E 1294-89, Standard Test Method for Pore Size Characteristics of Membrane Filters Using Automated Liquid Porosimeter by using automated bubble point method from ASTM Designation F 316 using a custom-built capillary flow porosimeter, in principle similar to a commercial apparatus from Porous Materials, Inc. (PMI), Ithaca, N.Y. Individual samples of 25 mm in diameter were wetted with isopropyl alcohol. Each sample was placed in a holder, and a differential pressure of air was applied and the fluid removed from the sample. The differential pressure at which wet flow is equal to one-half the dry flow (flow without wetting solvent) is used to calculate the mean flow pore size using supplied software.

(49) Flow Rate (also referred to as Flux) is the rate at which fluid passes through the sample of a given area and was measured by passing deionized water through filter medium samples having a diameter of 47 (9.6 cm.sup.2 filtration area) mm. The water was forced through the samples using hydraulic pressure (water head pressure) or pneumatic pressure (air pressure over water).

(50) The effective pore size of an electrospun mat can be measured using conventional membrane techniques such as bubble point, liquid-liquid porometry, and challenge test with particles of certain size. It is known that the effective pore size of a fibrous mat generally increases with the fiber diameter and decreases with porosity.

(51) Bubble point test provides a convenient way to measure effective pore size. It is calculated from the following equation:

(52) $P=\frac{2?}{r}\cos ?,$

(53) where P is the bubble point pressure, ? is the surface tension of the probe fluid, r is the pore radius, and ? is the liquid-solid contact angle.

(54) Membrane manufacturers assign nominal pore size ratings to commercial membrane filters, which are based on their retention characteristics.

(55) Retrovirus retention was tested following a Millipore test method. The bacteriophage PR772 challenge stream was prepared with a minimum titer of 1.0?10.sup.7 pfu/mL in a phosphate buffer saline (PBS) solution. Porous media to be tested were cut in 25 mm discs and sealed in overmolded polypropylene devices. These devices were then challenged by the above mentioned stream at 5 psi pressure after being wet by water at 25 psi pressure. The test was terminated after 100 ml of filtrate collection or after 4 hours of filtration, whichever came first. Quantification of bacteriophage in the initial and final feed were conducted on plates incubated overnight using a light box and a colony counter. Corresponding log retention values (LRV) were calculated.

(56) The following Examples of the present invention will demonstrate that an electrospun nanofiber mat can simultaneously possess both high permeability and high bacteria retention.

(57) Hereinafter the present invention will be described in more detail in the following examples. The invention will be further clarified by the following examples which are intended to be exemplary of the invention.

EXAMPLES

Example 1

(58) Nanofiber layers were made by electrospinning a solution of Nylon 6 polymer. Nylon 6 was supplied by BASF Corp., Florham Park, N.J., USA, under the trademark Ultramid B24. The spinning solution was prepared as a 20% Nylon stock solution in a mixture Acetic and Formic acids (2:1 weight ratio) for 5 hours at 80? C. and the stock solution was further diluted into 13 wt. % polymer solution and solvent ratio 2:2:1 of Formic:Acetic:Water, respectively, with the necessary formic acid and water additions. The viscosity of the resulting solution is about 100 cP. The solution was immediately spun using a 6-wire spinning electrode under 82 kV electric field. An antistatically-coated non-woven material was used to support the nanofiber mat during electrospinning. The average fiber diameter of electrospun mat produced was about 25-30 nm. The spin time was 30 minutes, after which the nanofiber layer was peeled off the mat, layered and placed in overmolded devices for retention testing.

(59) FIG. 2 depicts a frontal scanning electron micrograph of electrospun nylon fibers exemplified in Example 1.

(60) Table 1 displays a side by side comparison of two fully retrovirus retentive samples: one is a conventional immersion cast membrane (Retropore?) and the other is an electrospun nanofiber mat. Water permeability is reported for a 3 layer configuration for Retropore? and a 2 layer (30 minute spin time) configuration for the nanofiber media, since that is the corresponding number of layers typically needed to retain >6 logs of retrovirus retention.

(61) TABLE-US-00001 TABLE 1 Comparison of fully retentive (LRV > 6.2) samples in the device format; NFR membrane vs. electrospun nanofiber: Retropore? Electrospun membrane Nanofiber Mean flow IPA Bubble Point 122 132 (psi) Thickness (um) 438* 40** Porosity (%) 70 90 Water Permeability (LMH/psi) 35* 300** Retrovirus retention (LRV) >6.2* >6.2** *Measured on a three-layer membrane combination **Measured on a two-layer mat

(62) FIG. 4 depicts a graph of the bubble point distribution for the nylon fiber mat embodied in Example 1 and for a commercial retrovirus retentive membrane (Retropore?).

Example 2

(63) The procedure in Example 1 was followed. The spin time was varied from 5 to 60 minutes (FIG. 5.). PR772 retention was tested for a single layer of each nanofiber mat. Bacteriophage PR772 retention of the electrospun nanofiber mats increased with increasing spin time (increasing mat thickness).

(64) FIG. 5 is a graph of Log Reduction Value (LRV) of bacteriophage PR772 as a function of spin time of a nanofiber mat.

Example 3

(65) The procedure in Example 1 was followed, with solvent system for nylon being Formic acid/Water in weight ratio 4:1. The measured fiber diameter of produced electrospun mat was in the range of 25-30 nm, and the mean IPA bubble point in the range of 120-140 psi. 30 minute spin time samples were fully retrovirus retentive (LRV for bacteriophage PR772>6.2). FIG. 3 depicts a frontal scanning electron micrograph of the nylon fibers as exemplified in Example 3.

(66) Method of Use

(67) Electrospun nanofiber containing liquid filtration media, in accordance with the present invention are useful in the food, beverage, pharmaceuticals, biotechnology, microelectronics, chemical processing, water treatment, and other liquid treatment industries.

(68) Electrospun nanofiber containing liquid filtration media, in accordance with the present invention may be used for filtering, separating, identifying, and/or detecting microorganisms from a liquid sample or stream.

(69) Electrospun nanofiber containing liquid filtration media, in accordance with the present invention may be used with any liquid sample preparation methods including, but not limited to, chromatography; high pressure liquid chromatography (HPLC); electrophoresis; gel filtration; sample centrifugation; on-line sample preparation; diagnostic kits testing; diagnostic testing; high throughput screening; affinity binding assays; purification of a liquid sample; size-based separation of the components of the fluid sample; physical properties based separation of the components of the fluid sample; chemical properties based separation of the components of the fluid sample; biological properties based separation of the components of the fluid sample; electrostatic properties based separation of the components of the fluid sample; and, combinations thereof. Also, electrospun nanofiber containing liquid filtration media, in accordance with the present invention can be component or part of a larger device and/or system.

(70) Kits

(71) The invention also provides kits which may be used to remove microorganisms from a liquid sample. The kit may comprise, for example, one or more electrospun nanofiber containing liquid filtration medium in accordance with the present invention, as well as one or more liquid filtration devices, support or substrate for the medium. The kit may contain one or more controls, and may optionally include various buffers useful in the methods of practicing the invention, such as wash buffers for eliminating reagents or eliminating non-specifically retained or bound material may optionally be included in the kit.

(72) Other optional kit reagents include an elution buffer. Each of the buffers may be provided in a separate container as a solution. Alternatively the buffers may be provided in dry form or as a powder and may be made up as a solution according to the user's desired application. In this case the buffers may be provided in packets. The kit may provide a power source in instances where the device is automated as well as a means of providing an external force such as a vacuum pump. The kit may also include instructions for using the electrospun nanofiber containing liquid filtration medium, device, support or substrate, and/or for making up reagents suitable for use with the invention, and methods of practicing invention. Optional software for recording and analyzing data obtained while practicing the methods of the invention or while using the device of the invention may also be included.

(73) The term kit includes, for example, each of the components combined in a single package, the components individually packaged and sold together, or the components presented together in a catalog (e.g., on the same page or double-page spread in the catalog).

(74) The disclosure set forth above may encompass multiple distinct inventions with independent utility. Although each of these inventions has been disclosed in its preferred form(s), the specific embodiments thereof as disclosed and illustrated herein are not to be considered in a limiting sense, because numerous variations are possible. The subject matter of the inventions includes all novel and nonobvious combinations and subcombinations of the various elements, features, functions, and/or properties disclosed herein. The following claims particularly point out certain combinations and subcombinations regarded as novel and nonobvious. Inventions embodied in other combinations and subcombinations of features, functions, elements, and/or properties may be claimed in applications claiming priority from this or a related application. Such claims, whether directed to a different invention or to the same invention, and whether broader, narrower, equal, or different in scope to the original claims, also are regarded as included within the subject matter of the inventions of the present disclosure.